Biomedical subjects

Shigeru Kinoshita

Publications and source records attributed to Shigeru Kinoshita.

At least 37 records · Page 2Linked to original sources

PubMed2006

Cytomegalovirus in aqueous humor from an eye with corneal endotheliitis.

PURPOSE: To report cytomegalovirus (CMV) DNA in aqueous humor from a patient with unilateral corneal endotheliitis. DESIGN: Case report. METHODS: A 51-year-old man presented with unilateral corneal endotheliitis with linear keratic precipitates and coin-shaped lesions. Tear and aqueous humor samples were subjected to polymerase chain reaction to look for DNA from herpes simplex virus (HSV), varicella zoster virus (VZV), and CMV. RESULTS: Aqueous humor from the diseased eye contained DNA from CMV but not HSV or VZV. Its specificity was confirmed by Southern blot tests. Intravenous ganciclovir treatment resulted in the localization of his corneal edema and the reduction in keratic precipitates. There was severe destruction of corneal endothelial cells. CMV DNA was not detected in tears or control samples. CONCLUSIONS: In this healthy man with corneal endotheliitis, we detected CMV DNA in aqueous humor from the affected eye, but not HSV or VZV. This suggests that CMV may cause corneal endotheliitis in patients without immunodeficiency.

Acyclovir↗

PubMed2006

Histopathology of blepharoptosis induced by prolonged hard contact lens wear.

PURPOSE: To clarify histopathologically the structural features of blepharoptosis in prolonged hard contact lens wearers. DESIGN: Retrospective case-control study. METHODS: Biopsy specimens from identical sites at the levator aponeurosis and Mueller muscle from 15 long-term hard contact lens wearers were examined histopathologically (group 1). They comprised two men and 13 women with bilateral blepharoptosis ranging in age from 26 to 59 years (mean +/- SD, 44.4 +/- 10.70 years). The average length of hard contact lens wear was 25.4 years (range 12 to 40 years), and the average spherical equivalent refractive error was -9.100 diopters (range -2.825 to -20.375 diopters). We also examined specimens from 15 patients with involutional blepharoptosis who underwent levator resection; they comprised three men and 12 women ranging in age from 64 to 79 years (mean +/- SD, 72.3 +/- 4.38 years). RESULTS: All patients in group 1 manifested fibrosis and negligible fatty degeneration in Mueller muscle. In group 2, we detected mild fibrosis in Mueller muscle and fatty degeneration of the aponeurosis and Mueller muscle. CONCLUSIONS: Prolonged hard contact lens wear induces fibrosis in Mueller muscle and may result in contact lens-induced blepharoptosis.

Adult↗

PubMed2006

The features of dry eye disease in a Japanese elderly population.

PURPOSE: The purpose of this study is to assess the features of dry eye disease in a Japanese elderly population. METHODS: One hundred thirteen left eyes of 113 pensioners (50 males, 63 females; mean age, 67.5 +/- 5.7 years) aged over 60 years were recruited in this study. The subjects underwent careful slit-lamp examinations of the conjunctiva, ocular surface, and the eye lids. Tear film breakup time (BUT) examinations, Schirmer test-I, and fluorescein staining of the ocular surface and transillumination of the eyelids were also performed. Dry eye symptomatology was assessed with a symptom questionnaire. Japanese Dry Eye Diagnostic Criteria were used in this study. RESULTS: Ocular tiredness, irritation, dryness, and foreign body sensation were the most frequently reported symptoms by the patients. A total of 73.5% of the eyes had definite dry eyes. A total of 39.8% of the eyes had a Schirmer test reading <5 mm. Mean Schirmer test value was 9.4 +/- 7.8 mm. The mean BUT score was 4.0 +/- 2.8 seconds. A total of 76.9% of the eyes had positive fluorescein staining of the cornea. Meibomian gland dysfunction and conjunctivochalasis were found as frequent factors in relation to dry eye disease with meibomian grand dropout showing positive correlation with tear instability. CONCLUSION: Qualitative and quantitative disorders of the tear film were far more common than recognized in this population of elderly subjects, meibomian gland dysfunction being the most common associate of the tear film disorder and dry eye status. Conjunctivochalasis (conjunctival laxity), although commonly associated with dry eye disease in the elderly, was observed not to be related to age or gender in this study.

Age Factors↗

PubMed2006

Novel clinical application of sterilized, freeze-dried amniotic membrane to treat patients with pterygium.

PURPOSE: To evaluate the use of sterilized, freeze-dried amniotic membrane (FD-AM) transplantation for pterygium surgery. METHODS: This study involved a prospective, non-comparative, interventional case series. Thirteen eyes of 13 patients with primary (eight eyes) or recurrent (five eyes) pterygium were studied. After excision of the pterygium fibrous tissues and application of intraoperative use of mitomycin-C, sterilized FD-AM was sutured over the bare scleral defect. The integrity of the FD-AM graft, epithelialization over the FD-AM, pterygium recurrence and postoperative complications were evaluated. RESULTS: Postoperatively, the FD-AM was well retained in all patients, and complete epithelialization over the transplanted membrane was achieved within 1-2 weeks. All patients demonstrated early resolution of ocular inflammation and there was no recurrence of pterygium in any of the treated patients during the mean follow-up of 13.9 +/- 6.0 months. No ocular complications were noted following transplantation. CONCLUSION: Sterilized FD-AM showed excellent biocompatibility on the human ocular surface. This novel and promising biomaterial may be a useful alternative to conjunctival grafting in the treatment of pterygium.

Adult↗

PubMed2006

Clusters of corneal epithelial cells reside ectopically in human conjunctival epithelium.

PURPOSE: The ocular surface is covered by two biologically distinct epithelia: corneal and conjunctival. The expression of keratin12 (K12) is currently considered a hallmark of cornea-type differentiation. In the current study, the biological features of K12-positive cells in human bulbar conjunctival epithelium were examined. METHODS: Human conjunctival tissues were subjected to investigate the K12-positive cells in conjunctiva by immunostaining, in situ hybridization, Western blot analysis, reverse transcriptase-polymerase chain reaction (RT-PCR), and fluorescence-activated cell sorting (FACS). Gene expression profiling of these cells was performed with introduced amplified-fragment length polymorphism (iAFLP). To determine the presence of stem- or progenitor cells, immunostaining and colony-forming assays were performed. RESULTS: Western blot analysis, RT-PCR revealed that K12 was expressed in conjunctival epithelium. Immunostaining analysis showed that K12-positive cells reside mainly in clusters in conjunctival epithelium. FACS analysis showed that 0.2% to 1.7% of conjunctival epithelial cells collected from the inferior bulbar conjunctiva were K12 positive. iAFLP analysis revealed that the gene expression patterns of these cells were highly similar to that of corneal epithelial cells. p63 and ABCG2 were expressed beneath the K12-positive cells. Some colony-forming cells expressed K12. CONCLUSIONS: The K12-positive cells appear to be ectopically residing, self-maintaining corneal epithelial cells in the conjunctival epithelium.

Aged↗

PubMed2006

The use of autologous serum in the development of corneal and oral epithelial equivalents in patients with Stevens-Johnson syndrome.

PURPOSE: To evaluate the use of autologous serum (AS) from patients with severe ocular surface disease (OSD) in the development of transplantable corneal and oral epithelial tissue equivalents and to compare it with the use of conventional culture methods by using fetal bovine serum (FBS). METHODS: AS was obtained from patients with severe OSD secondary to Stevens-Johnson syndrome. Corneal and oral epithelial cells were cultivated in medium supplemented with either AS or FBS. Corneal and oral epithelial equivalents were constructed on denuded amniotic membranes. The bromodeoxyuridine (BrdU) ELISA cell proliferation assay and colony-forming efficiency (CFE) of cells cultivated in AS- or FBS-supplemented media were compared. The morphologic characteristics and the basement membrane assembly of cultivated epithelial equivalents were analyzed by light and electron microscopy, as well as by immunohistochemistry. RESULTS: BrdU proliferation assay and CFE analysis showed that human corneal and oral epithelial cells cultivated in AS-supplemented media had comparable proliferative capacities compared with FBS-supplemented media. The corneal and oral epithelial equivalents cultivated in AS- and FBS-supplemented media were morphologically similar and demonstrated the normal expression of tissue-specific keratins and basement membrane assembly. The presence of a well-formed stratified epithelium, a basement membrane, and hemidesmosomal attachments was confirmed by electron microscopy. CONCLUSIONS: AS-supplemented cultures were effective in supporting the proliferation of human corneal and oral epithelial cells, as well as the development of transplantable epithelial equivalents. The use of AS is of clinical importance in the development of autologous xenobiotic-free bioengineered ocular surface equivalents for clinical transplantation.

Adult↗

PubMed2006

Unique distribution of thrombospondin-1 in human ocular surface epithelium.

PURPOSE: The study was conducted to elucidate the detailed expression pattern of angiogenesis-related factors in human ocular surface epithelium. The focus was factors with significantly higher gene expression in corneal epithelium (CE) than in conjunctival epithelium (CJE). METHODS: The relative gene expression of 36 angiogenesis-related factors was compared in human CE and CJE, by using the introduced amplified fragment-length polymorphism (iAFLP) METHOD: Also examined were the expression patterns in the CE, limbal epithelium (LE), and CJE of factors with significantly higher expression in the CE, by using real-time PCR, in situ hybridization, immunohistochemistry, and immunoelectron microscopy. RESULTS: Only thrombospondin (TSP)-1 exhibited significantly higher expression in the CE. In situ hybridization and real-time PCR showed TSP-1 transcripts in the basal cells of the CE and LE. Compared with the CJE, they were significantly upregulated at those sites. Immunohistochemistry revealed that TSP-1 was strongly expressed in the basal region of the CE. Its expression was faint in LE and absent in CJE. Immunoelectron microscopy revealed that the CE and LE demonstrated TSP-1 labeling just below the epithelium, in the basal region of basal cells, and occasionally in the basal cell membrane. There was little or no labeling in the CJE. CONCLUSIONS: In the human ocular surface epithelium, basal cells of the CE and LE, but not of the CJE, synthesize TSP-1. High levels of TSP-1 are present only just below the CE. Its unique distribution may be related to corneal avascularity and integrity.

Angiogenic Proteins↗

PubMed2006

Establishment of a cultivated human conjunctival epithelium as an alternative tissue source for autologous corneal epithelial transplantation.

PURPOSE: The corneal epithelium is essential for maintaining corneal transparency, and efforts have been made to develop improved techniques for corneal epithelial transplantation in patients with total limbal failure. We evaluated the suitability of transplanted cultivated human conjunctival epithelium (HCjE) as a corneal epithelium replacement in rabbits with total corneal and limbal deficiency. METHODS: HCjE cells, cultivated on human amniotic membrane (AM) to confluence and exposed to an air-liquid interface (air-lifted), were transplanted onto denuded rabbit corneas and monitored for 2 weeks. The cultivated HCjE sheet and the engrafted epithelium were analyzed by immunohistochemistry and transmission electron microscopy (TEM). RESULTS: The transplanted HCjE remained transparent, smooth, and without epithelial defects during the follow-up period. Both the cultivated HCjE cells and the engrafted epithelium manifested five to six layers of stratified squamous epithelium similar in morphology to normal corneal epithelium. The basal cells expressed the putative stem cell markers (ABCG2 and P63) and hemidesmosome and desmosome component proteins. The cytokeratins (CK4, CK13, CK3, and CK12) and MUC4 were found in the engrafted epithelium. However, MUC5AC was not expressed. The results indicate that HCjE cultivated on AM has the potential to be used as an alternative corneal epithelium. CONCLUSIONS: The transplantation of cultivated HCjE sheets is a promising technique for the treatment of eyes with limbal failure.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMed2006

[A case of severe glaucoma with pseudopemphigoid successfully treated by filtration surgery using amniotic membrane].

BACKGROUND: It is necessary to decrease topical anti-glaucoma medication for severe glaucoma with pseudopemphigoid caused by anti-glaucoma eye drops. Glaucoma filtrating surgery is often needed instead of medication, but the prognosis is poor because it induces scar fomation and makes the filtrating bleb vanish. CASE: An 85-year-old male patient with exfoliation syndrome had twice undergone glaucoma surgery about ten years previously. His intra-ocular pressure (IOP) was high despite topical anti-glaucoma medication. At the first examination in our hospital, he had severe superficial punctate keratopathy, blephariticshortening and symblepharon, and we therefore diagnosed severe pseudopemphigoid induced by anti-glaucoma eye drops. Because his IOP could not be controlled by topical and general medication, we conducted a glaucoma filtrating operation using amniotic membrane. CONCLUSION: The administration of oral anti-inflammatory drugs before and after surgery and the use of amniotic membrane prevented post-operative scar formation and the progress of symblepharon, resulting in the successful control of IOP after surgery.

Aged, 80 and over↗

PubMed2006

[Relationship between compliance and background factors of glaucoma patients].

PURPOSE: To obtain useful information for the improvement of patient compliance in the medical treatment of glaucoma. SUBJECTS AND METHODS: An unsigned attitude survey on glaucoma and eye drops was conducted with 431 glaucoma patients. Correlation between compliance and individual(objective and subjective) factors was investigated. Objective factors were age, sex, number of eye drop products, duration of disease, and degree of visual field defects. Subjective factors were "stinging sensation with application of eye drops" and "worry about blindness". RESULTS: Thirty-eight percent of the glaucoma patients were compliant, instilling eye drops at the right time and with good technique. For the factor "duration of disease", patients who had glaucoma for 6-10 years had the best compliance. Compliance became poorer with an increasing number of eye drop products. In the subjective factors, less pain and worry led patients to better compliance. CONCLUSION: To improve compliance with medical treatment, we should categorize patients based on objective factors, eliminating their pain and worry, and instructing them not only in the frequency of instillation but also in the technique of administration.

Attitude↗

PubMed2005

Gene expression and immunolocalisation of a calcium-activated chloride channel during the stratification of cultivated and developing corneal epithelium.

The spatial and temporal localisation of a calcium-activated chloride channel (CLCA) and its mRNA was investigated, during the in vivo and in vitro development of stratified epithelia, by fluorescence immunohistochemistry and quantitative polymerase chain reaction in embryonic chicken corneas and the expansion of excised human corneal stem cells on amniotic membrane. Single-layered human epithelial cultures on amniotic membrane and early day embryonic chicken corneas expressed relatively little human CLCA2 or its chicken homologue. However, as the epithelium in both models matured and the number of cell-layers increased, the gene expression level and protein staining intensity increased, primarily within the basal cells of both the cultured and embryonic tissues. These results demonstrate that human CLCA2 protein and mRNA expression are elevated during epithelial stratification, suggesting that this protein plays a role in the growth of multi-layered corneal epithelia during both natural development and tissue cultivation.

Animals↗

PubMed2005

Pathological keratinisation in the conjunctival epithelium of Sjögren's syndrome.

Our previous gene expression analysis suggested that conjunctival epithelial cells of Sjögren's syndrome (SS) are inclined to hyper-proliferation and keratinisation status. The goal of this study is to elucidate whether such pathological situations really exist in the conjunctival epithelium of SS. Also, involvement of inflammatory cytokines in this disease was investigated. Conjunctival tissues or cells obtained from 12 SS patients and 13 normal subjects were subjected to indirect immunostaining to analyse expression of transglutaminase 1 (TGase1), involucrin, keratins 1, 4, 10 and 13. The number of proliferative cells was also analysed by immunostaining of Ki67 antigen. Additionally, changes in gene expression of TGase1 and involucrin after stimulation by IL-1 or IFN-gamma were quantified by real-time RT-PCR. TGase1 and involucrin were up-regulated in the conjunctival epithelium of SS patients. Although not statistically significant, Ki67 positive proliferative cells were slightly increased in SS patients. IFN-gamma stimulation significantly up-regulated TGase1 and unexpectedly repressed involucrin gene expression. IL-1 did not render any significant changes in the expression of these genes. These results suggest the existence of pathological keratinisation in the conjunctival epithelium of SS and also support our hypothesis that inflammatory cytokines may be involved in the ocular surface pathological changes in SS.

Aged↗

PubMed2005

Expression and tissue distribution of p63 isoforms in human ocular surface epithelia.

The functional significance of p63 in regulating cell proliferation in various stratified epithelial cells has previously been proposed. More than six isoforms have been reported for this protein; however, it is not yet clearly understood how functionally different these isoforms are. To investigate how these isoforms are used in ocular surface epithelia, we studied the spatial distribution of p63 isoforms within human ocular surface epithelia. Individual layers (basal, intermediate, and superficial) of the human ocular surface epithelia (cornea, limbus, and conjunctiva) were selectively obtained using a laser micro-dissection device. These samples were equally amplified and subjected to RT-PCR analysis with primer pairs, which specifically amplify each of five isoform-determining regions or each of six p63 isoforms. Regarding the N-terminal region, the TA domain was not detected in all samples, while a DeltaNp63 specific region was detected in the basal-intermediate region of all types of epithelia and in the superficial layer of the limbus. Regarding the C-terminal region, an alpha-isoform specific region was detected in all layers of the conjunctiva and limbus, as well as in the basal to intermediate layers of the cornea. A beta-isoform specific region was detected in the basal to intermediate layers of the limbus. A gamma-isoform specific region was detected in almost all layers of all epithelia. Among the six p63 isoforms, only DeltaNp63alpha was detected in the basal to intermediate layers of the limbus and conjunctiva. These results suggest that DeltaNp63alpha is the most dominant isoform within human ocular surface epithelia. This isoform may contribute, at least in part, to the maintenance of cell proliferative capacity within the ocular surface epithelia.

Aged↗

PubMed2005

Triggering of TLR3 by polyI:C in human corneal epithelial cells to induce inflammatory cytokines.

Epithelial cells of the ocular surface are key in the first-line defense as a part of the mucosal immune system against pathogens. We investigated whether polyI:C induces the production by human corneal epithelial cells (HCEC) of pro-inflammatory cytokines and IFN-beta, and whether Toll-like receptor (TLR)-3 expression is amplified by polyI:C. TLR3 was expressed on the surface of HCEC. Stimulation with polyI:C elicited the elevated production and mRNA expression of IL-6 and IL-8 in HCEC. While polyI:C induced IFN-beta, far stronger than human fibroblasts, and TLR3 gene expression in HCEC, LPS stimulation did not. Similarly, polyI:C, but not LPS, induced the gene expression of IkappaBalpha and MAIL, members of the IkappaB family, in HCEC. The innate immune response of HCEC is distinct from that of immune-competent cells, and we suggest that this is indicative of the symbiotic relationship between corneal epithelium and microbes inhabiting the ocular surface.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Allogeneic corneal tolerance in rodents with long-term graft survival.

BACKGROUND: Healthy C57BL/6 orthotopic corneal allografts in place for more than 8 weeks in BALB/c mice (acceptor8w+) can survive indefinitely due to active suppression of the donor-specific delayed-type hypersensitivity (DTH) response. This suggests a state of tolerance in the acceptor mice, however, the mechanism(s) underlying this acceptance remains to be demonstrated. We investigated the relationship between tolerance-induction and the DTH response using murine re-grafting models to explore the possibility of promoting allogeneic corneal regraft acceptance in high-risk graft beds. METHODS: Acceptor8w+ BALB/c mice received C57BL/6- or C3H corneal regrafts onto the same eye. Re-grafting models were prepared by inducing corneal neovascularization in the graft beds of naive BALB/c mice 2 weeks before corneal allografting. These mice were intravenously (iv) injected with purified splenic T cells or T-cell-depleted splenocytes from acceptor8w+ mice at the time they received re-grafts of C57BL/6 corneas. We also iv injected acceptor8w+ splenocytes into mice bearing healthy primary corneal allografts for 4 weeks (acceptor4w) and assessed their DTH response to C57BL/6 alloantigen(s). In those experiments, acceptor4w mice received a C57BL/6 corneal regraft onto the same eye. RESULTS: In all acceptor8w+ mice there was indefinite survival of C57BL/6-, but not of C3H regrafts. The iv injection of T cells, but not of T-cell-depleted populations, from acceptor8w+ splenocytes promoted allograft survival. Acceptor4w mice iv injected with acceptor8w+ splenocytes manifested a reduced C57BL/6-specific DTH response and the survival rate of C57BL/6 regrafts was increased from 0% to 87.5%. CONCLUSION: As donor-specific T cells from acceptor8w+ mice induced prolonged regraft survival, we posit that the active suppression of DTH responses by T cells may have contributed to indefinite allogeneic regraft survival via the induction of corneal allograft tolerance.

Adoptive Transfer↗

PubMed2005

Platelet aggregates in various stages of diabetic retinopathy: evaluation using the particle-counting light-scattering method.

AIMS: To investigate the possible correlation between platelet aggregation and the severity of diabetic retinopathy using the light-scattering method. METHODS: Using a light-scattering platelet aggregometer, we measured spontaneous platelet aggregation in 86 diabetics with retinopathy of varying severity and 30 healthy volunteers (controls). Platelet aggregates were classified as small, medium, and large according to their light intensity; patients were grouped based on the severity of retinopathy. In each patient group, we recorded for 10 min the total light intensities emitted by each aggregate size in the area under the curve (AUC). Then, we compared the AUC of each level of retinopathy severity with the controls and determined the correlation between the AUC of each aggregate size and each severity level. RESULTS: Of the 86 diabetics, 22 had no apparent retinopathy (NAR), 13 had mild nonproliferative diabetic retinopathy (NPDR), 17 had moderate NPDR, 12 had severe NPDR, and 22 had proliferative diabetic retinopathy (PDR). While the NAR group manifested significantly more small aggregates than the controls (20.5 x 10(6) versus 8.3 x 10(6) a.u., p=0.024), none of the other groups showed a significant increase in aggregates of any size. In the AUC of large aggregates, there was a weak-positive correlation with the severity of retinopathy (r=0.255, p=0.018); in the AUC of small and medium aggregates, there was no correlation. CONCLUSION: Although we did not find a significant correlation between platelet aggregation and the severity of diabetic retinopathy, our pilot study did detect some tendencies. Further studies on larger populations are underway to determine whether these tendencies are real.

Aged↗

PubMed2005

The quantification of hCLCA2 and colocalisation with integrin beta4 in stratified human epithelia.

Human calcium-activated chloride channel 2 (hCLCA2) belongs to a family of multifunctional proteins and is localised mainly in basal cells of squamous epithelia. However, its function is still not fully understood. Relative amounts of hCLCA2 were analysed using real-time PCR in several human epithelial tissues and tissues expressing high amounts were identified. These tissues then underwent double immunolabelling with anti-hCLCA2 antibodies and antibodies against the adhesion molecules integrin beta4 and collagen VII and were visualised by fluorescence microscopy. Real-time PCR found hCLCA2 gene expression to be primarily associated with stratified squamous epithelia. Subsequent immunohistochemistry clearly demonstrated colocalisation between hCLCA2 and integrin beta4. This study reports on a possible underlying relationship between hCLCA2 and stratified epithelia and the close association of hCLCA2 with basal cell adhesion molecules in normal tissue, suggesting it may play an important role in basal cell attachment in stratified epithelia.

Chloride Channels↗

PubMed2005

A new non-trocar system for 25-gauge transconjunctival pars plana vitrectomy.

PURPOSE: To introduce a new non-trocar system for 25-gauge transconjunctival pars plana vitrectomy (PPV). DESIGN: Study of a new surgical instrument. METHODS: This new non-trocar system for 25-gauge transconjunctival PPV consists of a contact lens ring featuring four projections containing 1.0-mm diameter holes located 3.0 mm from the ring edge, a wedge-shaped 25-gauge infusion cannula, and two plugs. RESULTS: The ring is fixed with 7-0 silk sutures at the 3- and 9-o'clock positions on the limbus. Using the 25-gauge needle, three conjunctival and scleral incisions (n = 3) are made at the projection holes located inferotemporally, superonasally, and superotemporally. No intra- or postoperative complications were encountered in 14 patients treated by this non-trocar 25-gauge transconjunctival PPV. CONCLUSION: Using the system introduced here, 25-gauge transconjunctival PPV can be performed without suturing sclerotomies and without intra- or postoperative complications.

Conjunctiva↗